Results of an Inpatient Preventive Health Care Program to Improve Quality of Life, Psychosocial Health, and Work Ability in Austria

Objective: The Austrian Federal Pension Insurance (PVA) developed a preventive inpatient health program, “Gesundheitsvorsorge-Aktiv (GVA),” for patients with musculoskeletal disorders. Individualized modular interventions and therapeutical measures (movement optimization, movement motivation, and mental health) are designed to improve occupational participation by influencing lifestyle factors and health-related quality of life. The study aimed to evaluate the new prevention-oriented and more personalized inpatient health program GVA.Methods: Patients underwent a standard inpatient health program, with emphasis on exercise management, exercise motivation, or psychological aspects. Submodule-dependent outcomes were assessed in patients (n = 330) at the start, end of treatment, and 6 months thereafter. Quality of Life (EQ-5D-5L), psychosocial aspects of the Patient Health Questionnaire (PHQ-D), and Work Ability Index (WAI) were queried.Results: The results consistently showed positive short and long-term effects. The subjective assessments of current work ability improved while the impairment of work performance was reduced. Positive changes in the psychosocial sphere were observed, alongside improvements in the health-related quality of life. Patients in the exercise optimization module performed better in all respects.Conclusion: In summary, GVA represents a valuable preventive health measure that leads to a holistic increase in well-being and can also ensure the maintenance of the ability to work

LAMTOR2-mediated modulation of NGF/MAPK activation kinetics during differentiation of PC12 cells.

LAMTOR2 (p14), a part of the larger LAMTOR/Ragulator complex, plays a crucial role in EGF-dependent activation of p42/44 mitogen-activated protein kinases (MAPK, ERK1/2). In this study, we investigated the role of LAMTOR2 in nerve growth factor (NGF)-mediated neuronal differentiation. Stimulation of PC12 (rat adrenal pheochromocytoma) cells with NGF is known to activate the MAPK. Pharmacological inhibition of MEK1 as well as siRNA-mediated knockdown of both p42 and p44 MAPK resulted in inhibition of neurite outgrowth. Contrary to expectations, siRNA-mediated knockdown of LAMTOR2 effectively augmented neurite formation and neurite length of PC12 cells. Ectopic expression of a siRNA-resistant LAMTOR2 ortholog reversed this phenotype back to wildtype levels, ruling out nonspecific off-target effects of this LAMTOR2 siRNA approach. Mechanistically, LAMTOR2 siRNA treatment significantly enhanced NGF-dependent MAPK activity, and this effect again was reversed upon expression of the siRNA-resistant LAMTOR2 ortholog. Studies of intracellular trafficking of the NGF receptor TrkA revealed a rapid colocalization with early endosomes, which was modulated by LAMTOR2 siRNA. Inhibition of LAMTOR2 and concomitant destabilization of the remaining members of the LAMTOR complex apparently leads to a faster release of the TrkA/MAPK signaling module and nuclear increase of activated MAPK. These results suggest a modulatory role of the MEK1 adapter protein LAMTOR2 in NGF-mediated MAPK activation required for induction of neurite outgrowth in PC12 cells

LAMTOR2 is involved in endosomal trafficking.

<p>(A, B) PC12 cells were transfected with control or specific siRNA for LAMTOR2. After 24 h, cells were stimulated with NGF (200 ng/ml) for various time points (0 = control, or 5 and 15 min), fixed and stained for EEA1 (early endosomes), pan-TrkA and Hoechst (nucleus). EEA 1 is shown in red and pan-TrkA in green. Colocalization of EEA1 and pan-TrkA is shown in yellow (arrows). Scale bar = 10 µm. Colocalization tests were performed as described in material and methods section. Colocalization in LAMTOR2 knockdown cells is expressed as fold of control (f.o.c.; control values in control siRNA transfected cells: 0 min 20.99±8.55%, 5 min 22.74±7.94%, 15 min 24.94±9.04%). Values represents the means ± SEM, n = 3–5. Differences were analyzed using unpaired two-tailed t-test *p<0.05.</p

LAMTOR2 phenotype is rescued with a human LAMTOR2 ortholog.

<p>(A, B) Control or LAMTOR2 siRNA silenced cells were cotransfected with a human LAMTOR2 ortholog and stimulated with NGF (25ng/ml). Pictures of 3–5 independent microscopic fields were taken after 16 h (A) and neurite-bearing cells were calculated as fold of control (f.o.c. control value was 22.45±1.39% neurite-bearing cells) in (B). Pictures are representative of 4 independent experiments. Values represent the mean ± SEM, n = 4. Scale bar = 20 µm. Differences were analyzed using unpaired two-tailed t-test: ***p<0.001.</p

NGF-induced neurite formation depends on the activation of p42/44 MAPK.

<p>(A) Cells left untreated (co), stimulated with NGF (25 ng/ml) or alternatively with NGF (25 ng/ml) and PD098059 (50 µM) for 16 h and were stained with Phalloidin-TRITC and Hoechst 33342. Pictures were taken of 3–4 independent microscopic fields and are representative of 3 experiments. Scale bar = 20 µm. (B, C) PC12 cells were stimulated with either NGF (25 ng/ml) or EGF (100 ng/ml) and activation of p42/44 MAPK was measured by Bio-Plex phosphoprotein detection assay at various time points (0 min –20 min). Differences were analyzed using one-way ANOVA followed by Dunnett’s multiple comparison test: *p<0.05, **p<0.01, ***p<0.001 (B). Total cell lysates were analyzed with western blotting done in parallel for 0 and 5 min. (D, E) PC12 cells were transfected with control or specific siRNA for MAPK. After 72 h in culture, cells were stimulated with NGF (5 ng/ml). Additional 3 d pictures were taken of 3–5 microscopic fields (D) and neurite-bearing cells are expressed as fold of control (f.o.c.; control value was 2.16±0.53% neurite-bearing cells) in (E). Pictures are representative of 5 independent experiments. Values represent the mean ± SEM, n = 5. Scale bar = 20 µm. Differences were analyzed using one-way ANOVA followed by Dunnett’s multiple comparison test: *p<0.05, **p<0.01.</p

SiRNA-mediated knockdown of LAMTOR2 scaffold protein leads to increased NGF- mediated MAPK activation.

<p>(A, B) PC12 cells were transfected with control or specific siRNA for LAMTOR2. After 24 h, cells were left untreated (co) or stimulated with either NGF (25 ng/ml) or EGF (100 ng/ml). After various time points (0, 2, 5, 10, 20 min), activation of p42/44 MAPK was measured with the Bio-Plex phosphoprotein detection assay. Values represent the mean ± SEM, n = 3. Differences were analyzed using unpaired two-tailed t-test: ***p<0.001. (B) In parallel, after 5 min, total cell lysates were analyzed by western blotting technique. (C) Cells were transfected with LAMTOR2 siRNA plus control vector or plus a human LAMTOR2 ortholog. After stimulation with NGF (25 ng/ml) for 5 min Bio-Plex phosphoprotein detection assay was carried out and p42/44 MAPK activation expressed as fold of control (f.o.c. control value was 4.57±0.67%). Values represent the mean ± SEM, n = 7. Differences were analyzed using unpaired two-tailed t-test: *p<0.05.</p

LAMTOR2 adapter protein modulates NGF-mediated differentiation.

<p>(A, B) PC12 cells were stimulated with either NGF (25 ng/ml) or EGF (100 ng/ml). Neurite outgrowth was examined in 3–5 independent microscopic fields (72 h after stimulation shown in (A)). Percentages of neurite outgrowth (numbers reflecting neurite-bearing cells) after various time points (6 h–72 h) were calculated, counting 500–700 cells for each time point, and are depicted in (B). Microscopic images are representative of 3 independent experiments. Values represent the mean ± SEM, n = 3 (number of experiments). Scale bar = 20 µm. Differences were analyzed using one-way ANOVA followed by Dunnett’s multiple comparison test: ***p<0.001. (C) PC12 cells were transfected with control or specific LAMTOR2 siRNA, and mRNA expression levels were examined after 24 h, 48 h and 72 h by quantitative real-time RT-PCR; values represent the mean ± SEM, n = 3–5. Differences were analyzed using unpaired two-tailed t-test: ***p<0.001. (D) LAMTOR2 protein expression levels were shown by western blotting technique 48 h after transfection. Blot is representative of 3 independent experiments. (E) PC12 cells transfected with control or specific siRNA for LAMTOR2 were analyzed for protein expression of the other members of the LAMTOR complex, LAMTOR1 (p18), LAMTOR3 (MP1), LAMTOR4 (C7orf59) and LAMTOR5 (HBXIP) at 48 hours. Blots are representative of 3 experiments. (F) LAMTOR downregulation (%) was quantified and differences analyzed using unpaired two-tailed t-test: LAMTOR1 (p18; 7.63±1.50% reduction, **p<0.01), LAMTOR2 (p14; 60.25±13.54% reduction, *p<0.05), LAMTOR3 (MP1; 55.91±14.68% reduction, *p<0.05), LAMTOR4 (C7orf59; 51.85±12.52% reduction, *p<0.05) and LAMTOR5 (HBXIP; 41.06±10.33% reduction, *p<0.05).</p